Intrinsically asymmetric cell division plays an important role in development by contributing to cellular diversity and embryonic pattern formation. To understand asymmetric cell divisions it is necessary to understand the generation and maintenance of cell polarity. Elucidating mechanisms for the establishment and maintenance of cell polarity is an important goal not only because of the role of polarity in asymmetric cell division but also because cell polarity is the foundation for the proper development and function of most tissue and organ systems. We propose to continue our ongoing analysis of the generation of cell polarity during the asymmetric divisions of the model organism Caenorhabditis elegans. We will focus on the role of the PAR proteins in establishing cell polarity during the asymmetric divisions of the early embryo. PAR proteins are evolutionary conserved regulators of cell polarity with roles in a variety of cell types including polarized epithelial cells, neurons, oocytes and migratory cells of many types. We will carry out three related studies. In Aim 1 we will investigate the means by which the PAR proteins become asymmetrically distributed in the one-cell embryo by determining mechanisms of regulation of contractility of the actomyosin system in response to a polarity cue. In Aim 2 we will dissect the molecular mechanism of action of the PAR-3 protein by testing the function and localization of engineered mutant forms of the protein in living worms. In Aim 3 we will use RNA interference in two different approaches to identify substrates of the protein kinases PAR-1 and PAR-4 and then determine the mode of action of proteins we identify. We expect the results of our studies to contribute to our understanding of how cell polarity is regulated during embryonic development of C. elegans and humans.